The Protective Effect of Virus Capsids on RNA and DNA Virus Genomes in Wastewater

ABSTRACT Virus concentrations measured in municipal wastewater help inform both the water treatment necessary to protect human health and wastewater-based epidemiology. Wastewater measurements are typically PCR-based, and interpreting gene copy concentrations requires an understanding of the form and stability of the nucleic acids. Here, we study the persistence of model virus genomes in wastewater, the protective effects provided by the virus capsids, and the relative decay rates of genome and infectious viruses. In benchtop batch experiments at 25 °C, extraviral (+)ssRNA and dsDNA amplicons degraded by 90% within 15-19 minutes and 1.6-1.9 hours, respectively. When encapsidated, the T 90 for MS2 (+)ssRNA increased by 424× and the T 90 for T4 dsDNA increased by 52×. The (+)ssRNA decay rates were similar for a range of amplicon sizes. For our model phages MS2 and T4, the nucleic acid signal in untreated wastewater disappeared shortly after the viruses lost infectivity. Combined, these results suggest that most viral genome copies measured in wastewater are part of intact virus particles, that measured concentrations are independent of assay amplicon sizes, and that the virus genome decay rates of naked viruses are similar to inactivation rates. These findings will be valuable for the interpretation of wastewater virus measurements.